One of the challenges to the application of stem cells as therapeutics in regenerative medicine is the status of the microenvironment to which the stem cell therapy will be applied. Unfortunately, these specialized environments are frequently damaged or degenerated with age or insult. A damaged environment may not support stem cell repopulation following therapy, or may induce aberrant cellular function resulting in undesirable pathologies. Restorative efforts become even more complicated in multifunctional organs with cells having multiple phenotypes, such as female ovaries. Ovaries are dynamic organs with multiple cellular phenotypes derived from multiple lineages. For example, the gonad itself is generated from a population of somatic cells from the embryonic genital ridge, whereas germ cells are of extra-gonadal origin, migrating to the newly formed gonad shortly after its formation. In the neonate, surviving germ cells in the ovaries are arrested in the late diplotene stage of meiosis and enclosed by pre-granulosa cells of somatic cell linage in structures termed follicles. As such, regenerative efforts directed at rejuvenating female fertility or ovarian function in the aged or damaged ovary by targeting only one cell type may have limited success, due to the necessity to restore multiple cellular phenotypes. The goal of this study is to restore a healthy ovarian environment in the aged or damaged ovary through the generation of specialized somatic cells derived from embryonic stem (ES) and induced pluripotent stem (iPS) cells. Using ES and IPS cells derived from TgOG2 transgenic mice with germline-specific expression of GFP, transduced with a somatic cell-specific expression of DsRed, both somatic and germ cell populations can be distinguished in vitro. Specifically, Aim I will identify and characterize ovarian specific somatic cells in vitro, while Aim II will use transplantation of isolated germline and somatic cell populations derived from ES and iPS cells into aged or chemically damaged ovaries to restore follicle formation in vivo. Using a marker specific for ovarian somatic cells, F0XL2 (a FOX winged-helix forkhead gene transcription factor), a sub-population of differentiated ES and iPS cells can be identified and isolated. These isolated somatic cells can then be transplanted into follicle depleted adult mouse ovaries. Somatic cells are required for follicle formation, and ultimately steroid hormone production and ovulation, thus comprising a critical component of the ovarian environment. Increasing the healthy somatic cell population in the aged or damaged ovary represents an eariy, yet crucial, step in restoration of ovarian function. It is thought that by restoring this environment, which is damaged by. aging, that ovarian function can be prolonged or restored, ultimately alleviating the menopause.